Wound healing composition and method for use thereof

ABSTRACT

A method for reducing or preventing the formation of scar tissue is disclosed. The method comprises applying a pharmacologically effective amount of a composition comprising about 5.0% by weight of ovalbumin, about 1.0% phenoxyethanol, about 0.5% carbomer, and about 0.3% triethanolamine. A method for stimulating fibroblast production is also disclosed. The method comprises applying a pharmacologically effective amount of a composition comprising about 5.0% by weight of ovalbumin, about 1.0% phenoxyethanol, about 0.5% carbomer, and about 0.3% triethanolamine.

FIELD OF THE INVENTION

[0001] This invention pertains to wound healing compositions and,particularly, to such a composition comprising of a growth hormone andgrowth factor in a balanced mixture that will deliver the maximumtherapeutic results with non-healing wounds, burns, trauma, and certaindermatological disorders.

BACKGROUND OF THE INVENTION

[0002] 1. Overview of the Invention

[0003] There have been recent dramatic strides with the discovery ofgrowth factors as wound healing agents. Indeed, the discovery of growthfactors has triggered great optimism into the possibility of masteringthe art of wound healing and intense effort has been launched on thepart of medical researchers and pharmaceutical companies to procure,characterize, and harvest these healing enhancement agents. Because itis believed that administering growth factors to patients with dermaland subdermal wounds enhances the speed by which wounds heal, asdiscussed in greater detail below, it is the object of this invention toprovide a topical composition containing growth factors to increase thebody's wound healing properties.

[0004] 2. Background and Prior Art

[0005] Since their discovery almost 30 years ago, growth factors havebeen shown to stimulate neovascularization in vitro and in animalstudies. In the past 10 years, knowledge of growth factors has grownimmensely. Broadly defined, growth factors are multifunctional, locallyacting, intercellular signaling polypeptides which, among other things,organize and coordinate cellular profileration. Most growth factors arelarge peptides or glycoproteins secreted by many cells as a basefunction, or in response to a challenge, such as a wound or carcinogen.These peptides represent a system of signals that mediate physiologicand pathologic cellular growth and repair, including embryogenesis,wound healing and carcinogenesis.

[0006] Currently, there are two known classes of intercellular signalingproteins: (i) endocrine proteins which are long range signaling proteinsreleased into the circulation or other body fluids, and (ii) paracrineproteins which are short range signaling proteins that act locallywithin tissues. Growth factors are generally considered as paracrineproteins since they are predominantly short range locally acting,intercellular signaling proteins.

[0007] There exist six known varieties of growth factors:platelet-derived growth factor (PDFG), epidermal growth factor (EGF),fibroblast growth factor (FGF), insulin-like growth factor (IGF),transforming growth factor beta (TGFβ), and TGFβ Superfamily. Inaddition to their local mode of action, the different varieties ofgrowth factors share common biological properties. For example, growthfactor action is mediated by association with specific, high affinityreceptors expressed by the target cells, and growth factors are able toexert their biological effect at low concentrations (10⁻⁹ to 10⁻¹¹M).

[0008] Furthermore, on the cellular level, growth factors bind withligands on the cell surface to generate an intracellular signal on theinside of the cell to modify cellular behavior. Growth factors functionby binding to specific cell-surface receptors that are composed of threedistinct regions or domains: an extracellular domain, which binds to thegrowth factor ligand; a transmembrane domain; and an intracellulardomain. When the ligand binds to neighboring receptors, conformationalchanges occur that are transmitted to the intracellular domain andelicit a series of cytoplasmic changes leading to the initiation of thenucleic acid transcription. In many cases, these intracytoplasmic eventsare enacted through an enzyme, tyrosine kinase, that phosphorylatescytoplasmic proteins, some of which remain in the cytoplasm and somepass to the nucleus with a corresponding gene activation.

[0009] Studies have shown that fluid factors isolated from chronicwounds lack the presence of growth factors. Moreover, in vitro studieshave shown that growth factors added to wound fluid extracted frompostoperative or traumatic wounds have accelerated the wound healingcascade. These facts strongly suggest the important role of growthfactors.

[0010] One in vitro study examined the effect of human wound fluid onthe growth of human dermal fibroblasts and unbilical vein endothelialcells. Katz M H et al., J. AM. ACAD. DERMATOL., Human Wound Fluid fromAcute Wounds Stimulates Fibroblast and Endothelial Cell Growth,25:1054-1058 (December 1988). Katz el al. collected wound fluid from sixpatients undergoing split-thickness skin and wound fluid frompostoperative patients. After seeding the wound fluid in optimal growthmedia (control) on day 0, cultures of human dermal fibroblasts andumbilical vein endothelial cells were supplemented with or without acutewound fluid on days 1 and 3. The study found that 2% acute wound fluidstimulated the growth of human dermal fibroblasts and umbilical veinendothelial cells when these cells were cultured in 2% fetal bovineserum and endothelial growth medium, respectively. Wound fluid from thepostoperative patients caused the same level of stimulation.Furthermore, when anti-platelet-derived growth factor antibody was addedto wound fluid, there was a 45% mean reduction in the stimulatory effecton fibroblast growth. This result further suggests that platelet-derivedgrowth factor contributes to the fibroblast growth effect.

[0011] It has also been suggested that ulcer healing may be improved bythe exogenous provision of specific growth factors. Research has shownthat fibroblasts isolated from wound sites on patients proliferated at aslower rate and are morphologically distinct (larger and polyglonal inshape) from normal fibroblasts cells. Stanley A C et al., J. VASC.SURG., Reduced Growth of Dermal Fibroblasts from Chronic Venous Ulcerscan be Stimulated with Growth Factors, 26(6):994-999 (December 1997). Itwas also found that the decreased growth of wound fibroblasts werestimulated by growth factors FGF, EGF, IL-1.

[0012] A combination of growth factors produce a magnified effect instimulating protein synthesis and decreasing protein degradation. Theyhave also been shown to have additive effect in improving whole-body andmuscle kinetics. For example, it has been shown that the addition ofpure PDGF to a wound site involving the epidermis and dermis has littleeffect on the morphology or biochemistry of wound healing. Lynch S E etal., PROC. NAT'L. ACAD. SCI. USA, Role of Platelet-Derived Growth Factorin Wound Healing: Synergistic Effects with Other Growth Factors, 84(21):7696-7700 (November 1987). In contrast, the addition of partiallypurified PDGF results in significant dose-dependent increases in thewidth of the newly synthesized connective tissue and epidermal layers.Further, the addition of partially purified PDGF results in significantincreases in the rate of protein and DNA synthesis and the total contentof these components in biopsies taken from the wound site. Similareffects were obtained when IGF was added in combination with pure PDGF.Combining the growth factors caused a 2.4 fold increase in the width ofnewly formed connective tissue layer and a 95% increase in epidermalthickness compared with controls. Id. On the other hand, IGF appliedalone did not cause similar morphological changes, thus indicating thatthe synergistic actions of other factors with PDGF are important in themodulation of the wound healing process. Therefore, a combination ofgrowth factors and growth hormones have a potential amplified effect onaccelerated non-healing of ulcers, open wound, osteomyelitis, skeletalmuscle injuries such as sports and trauma, and burns and dermatologicaldisorders.

[0013] Animal and clinical trials using growth factor therapy haveproduced outstanding results. Healing of a variety of wounds in animalsand patients was enhanced by treatment with EGF or TGF-alpha. Severaldifferent studies of topically applied growth factors have shown toaccelerate healing by stimulating granulation tissue formation andenhancing epithelialization. Epidermal regeneration of partial thicknessburns on pigs or dermatome wounds on patients was accelerated withtopical application of EGF or TGF-alpha, and EGF treatment acceleratedhealing of gastroduodenal ulcers. Schultz G. et al., J. CELL. BIOCHEM.,EGF and TGF-Alpha in Wound Healing and Repair, 45(4):346-352 (April1991). EGF also increased tensile strength of skin incisions in rats andcorneal incisions in rabbits, cats, and primates. Id.

[0014] Specific use of particular factors is becoming an accepted modeof therapy for many patients with non-healing wounds. Accordingly, thereis ample scientific and clinical evidence suggesting that the biologicalactions of growth factors regulate cell division, differentiation,migrations and gene expression.

[0015] The present invention is a composition comprising a mixture ofgrowth factors and growth hormones in a dermatologically safe vehicle,such as a liquid, gel or cream. It is therefore an object of the presentinvention to provide a wound-healing composition comprising a proteingrowth factor to treat healing resistant wounds, dialetic ulcers,bedsores, burns, osteomyelitis, trauma wounds, subcutaneous trauma andvarious forms of dermatitis. The composition is also useful in reducingthe formation of scar tissue.

SUMMARY OF THE INVENTION

[0016] In accordance with the present invention, it has now beendiscovered that wounds may be effectively treated with the growthfactors that are contained in egg whites of hens. Specifically,topically administered growth factors in pharmacologically effectiveamounts are useful for increasing the local activity of the body's woundhealing process. Accordingly, the present invention is a low cost,over-the-counter, composition containing growth factors isolated from anaturally occurring source—egg whites. The composition may be used totreat any disease state where growth factors play a role includingdiabetic ulcers, nonhealing wounds, burns, osteomyelitis, trauma wounds,subcutaneous trauma and various forms of dermatitis.

[0017] In particular, the wound-healing composition is comprised ofphenoxyethanol, carbomer, protein and triethanolamine. The protein isovalbumin from egg whites, which includes a combination growth factorsincluding epidermal growth factor (EGF), transforming growth factoralpha (TGF-Alpha), fibroblast growth factor (acidic) (FGF-a), fibroblastgrowth factor (basic) (FGF-b).

[0018] In a preferred embodiment of the invention, the wound-healingcomposition comprises about 1.0% by weight of phenoxyethanol, about 0.5%carbomer, about 20% ovalbumin and about 0.3% triethanolamine. Thesecompounds are blended to produce a eutectic mixture in gel, lotion,cream or ointment form. The inventive composition can also be in powderform as well.

[0019] It is therefore an object of the present invention to treatmammalian nonhealing wounds manifested by diabetes, burns, trauma andsubcutaneous trauma, osteomyelitis, various surgical procedures, andvarious forms of dermatitis.

[0020] It is another object of the present invention to treat woundswith a topical agent that is easily administered, economical andwell-tolerated by patients.

[0021] It is another object of this invention to provide a compositionto stimulate fibroblast production for tissue repair.

[0022] It is yet another object of this invention to provide acomposition for the stimulation of hair follicle growth.

[0023] It is still another object of the present invention to provide acomposition for treating non healing wounds by the addition of anindividual or combination of growth factors including IGF, PDGF, FGF,EGF, TGFβ and TGFβ Superfamily.

[0024] These and other features and advantages of the present inventionwill be found in the following description of the preferred embodimentsand in the claims.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0025] Before the present composition for wound healing is disclosed anddescribed, it is to be understood that this invention is not limited tothe particular configurations, process steps, and materials disclosedherein as such configurations, process steps, and materials may varysomewhat. It is also to be understood that the terminology employedherein is used for the purpose of describing particular embodiments onlyand is not intended to be limiting since the scope of the presentinvention will be limited only by the appended claims and equivalentsthereof.

[0026] As described above, the present invention is directed to acomposition for stimulating cell growth and the healing of wounds byadministering a pharmacologically effective amount of grown factorsdirectly to the affected tissue. The composition may be used to treatany type of non-healing wound whereby fibroblast activity is reduced orineffective. Such wounds include diabetic non-healing wounds, burns,osteomyelitis, trauma wounds, subcutaneous trauma and various forms ofdermatitis. The composition may also be employed to stimulate hairgrowth on the scalp and other body areas.

[0027] The growth factors of the present invention have been identifiedin hen's egg whites. Ovalbumin is the major constituent of egg whitesfrom the hen comprising about 75% by weight of the egg white. Themolecular weight of ovalbumin is approximately 4,500,and ovalbumin isproduced under hormonal control by the bird oviduct. It may be isolatedand crystallized readily from the filtrate of an acidified mixture ofegg white and an equal volume of saturated ammonium sulfate. Sorensen etal., C. R. TRAV. LAB. CARLSBERG 12, 12 (1917). Alternative methods ofisolation of ovalbumin are disclosed by Kekwick et al., BIOCHEM JOURNAL30:227 (1930). Ovalbumin can also be separated by electrophoresis andchromatography from about 10 other minor components found in egg whitesincluding avidin (qv), lysozyme (qv), conalbumin (qv), and ovomucoid.

[0028] The structure of ovalbumin is that of a complex proteinconsisting of a single polypeptide chain of about 460 residues (abouthalf of which are hydrophobic), a maximum of 2 phosphate residues permole, and a oligosaccharide side chain composed of only mannose andglucosamine residues. See Narita, J. BIOCHEM. 52:367 (1962); Thompson etal., AUST. J. BIOL. SCIENCE 24: 525 (1971).

[0029] Ovalbumin is soluble in electrolyte free water and combines withsalts, acids and bases. Denaturation can be induced by heating to 56°C., by vigorous shaking, by electric current and by various chemicalssuch as acids, ammonium salts, heavy metal salts and alcohols. Suchmethods produce complete and irreversible denaturation. The isoelectricpoint of ovalbumin is 4.63. See Merck Index (12^(th) Ed. 1996).

[0030] In accordance with the compositions and method of the presentinvention, ovalbumin may be administered in the form of a pharmaceuticalcomposition additionally comprising pharmaceutically acceptable carrier.One skilled in the art will appreciate that suitable methods ofadministering the ovalbumin compositions to an animal, such as a mammal,are available and, although more than one method can be used toadminister a particular composition, a particular method and dosage canprovide a more immediate and more effective reaction than others.Pharmaceutically acceptable carriers are also well known to thoseskilled in the art. The choice of carrier will be determined, in part,both by the particular composition and by the particular method used toadminister the composition. Accordingly, there is a wide variety ofsuitable formulations of the pharmaceutical compositions of the presentinvention.

[0031] The present invention is preferable in the form of a topicaldosage form such as creams, ointments, lotions, gels or powders.

[0032] The present invention may be formulated as necessary withadditives used commonly in the pharmaceutical sciences, such assurfactants, oils and fats, polyhydric alcohols, lower alcohols,thickening agents, UV absorbents, light scattering agents,preservatives, antioxidants, antibiotics, chelating agents, pHregulators, flavoring agents, pigments and water.

[0033] Examples of surfactants include polyoxyethylene (hereinafterabbreviated as POE-) branched alkyl ethers such as POE-octyldodecylalcohol and POE-2-decyltetradecyl alcohol, POE-alkyl ethers such asPOE-oleyl alcohol ether and POE-cetyl alcohol ether, sorbitan esterssuch as sorbitan monooleate, sorbitan monoisostearate and sorbitanmonolaurate, POE-sorbitan esters such as POE-sorbitan monooleate,POE-sorbitan monoisostearate and POE-sorbitan monolaurate, fatty acidesters of glycerol such as glyceryl monooleate, glyceryl monostearateand glyceryl monomyristate, POE-fatty acid esters of glycerol such asPOE-glyceryl monooleate, POE-glyceryl monostearate and POE-glycerylmonomyristate, POE-dihydrocholesterol ester, POE-hardened castor oil,POE-hardened castor oil fatty acid esters such as POE-hardened castoroil isostearate, POE-alkylaryl ethers such as POE-octylphenol ether,glycerol esters such as glycerol monoisostearate and glycerolmonomyristate, POE-glycerol ethers such as POE-glycerol monoisostearateand POE-glycerol monomyristate, polyglycerol fatty acid esters such asdiglyceryl monostearate, decaglyceryl decastearate, decaglyceryldecaisostearate and diglyceryl diisostearte and other nonionicsurfactants; potassium salts, sodium salts, diethanolamine salts,triethanolamine salts, amino acid salts and other salts of higher fattyacids such as myristic acid, stearic acid, palmitic acid, behenic acid,isostearic acid and oleic acid, the above alkali salts of ethercarboxylic acids, salts of N-acylamino acids, N-acylsalconates, higheralkylsulfonates and other anionic surfactants; alkylamine salts,polyamine, aminoalcohol fatty acids, organic silicone resin, alkylquaternary ammonium salts and other cationic surfactants; and lecithin,betaine derivatives and other amphoteric surfactants.

[0034] Examples of oils and fats include vegetable oils and fats such ascastor-oil, olive oil, cacao oil, camellia oil, coconut oil, wood wax,jojoba oil, grape seed oil and avocado oil; animal oils and fats such asmink oil and egg yolk oil; waxes such as beeswax, whale wax, lanolin,carnauba wax and candelilla wax; hydrocarbons such as liquid paraffin,squalene, microcrystalline wax, ceresine wax; paraffin wax and vaseline;natural or synthetic fatty acids such as lauric acid, myristic acid,stearic acid, oleic acid, isostearic acid and behenic acid; natural orhigher alcohols such as cetanol, stearyl alcohol, hexyldecanol,octyldecanol and lauryl alcohol; and esters such as isopropyl myristate,isopropyl palmitate, octyldodecyl myristate, octyldodecyl oleate andcholesterol oleate.

[0035] Examples of polyhydric alcohols include ethylene glycol,polyethylene glycol, propylene glycol, 1,3-butyrene glycol, 1,4-butyreneglycol, diprophylene glycol, glycerol, diglycerol, triglycerol,tetraglycerol and other polyglycerols, glucose, maltose, maltitose,sucrose, fructose, xylitose, sorbitol, maltotriose, threitol anderythritol.

[0036] Examples of thickening agents include naturally-occurring highmolecular substances such as sodium alginate, xanthene gum, aluminumsilicate, quince seed extract, gum tragacanth, starch, collagen andsodium hyaluronate; semi-synthetic high molecular substances such asmethyl cellulose, hydroxyethyl cellulose, carboxymethyl cellulose,soluble starch and cationized cellulose; and synthetic high molecularsubstances such as carboxyvinyl polymer and polyvinyl alcohol.

[0037] Examples of UV absorbents include p-amnobenzoic acid,2-ethoxyethyl p-methoxycinnamate, isopropyl p-methoxycinnamate,butylmethoxybenzoylmethane,glyceryl-mono-2-ethylhexanoyl-di-p-methoxybenzophenone, digalloyltrileate, 2,2′-dihydroxy-4-methoxybenzophenone,ethyl-4-bishydroxypropylaminobenzoate,2-ethylhexyl-2-cyano-3,3′-diphenyl acrylate, ethylhexylp-methoxycinnamate, 2-ethylhexyl salicylate, glyceryl p-aminobenzoate,homomethyl salicylate, methyl o-aminobenzoate,2-hydroxy-4-methoxybenzophenone, amyl p-dimethylaminobenzoate,2-phenylbenzoimidazole-5-sulfonic acid and2-hydroxy-4-methoxybenzophenone-5-sulfonic acid.

[0038] Examples of preservatives include benzoates, salicylates,sorbates, dehydroacetates, p-oxybenzoates,2,4,4′-trichloro-2′-hydroxydiphenyl ether, 3,4,4′-trichlorocarbanilide,benzalkonium chloride, hinokitiol, resorcinol and ethanol.

[0039] Examples of antioxidants include tocopherol, ascorbic acid,butylhydroxyanisole, dibutylhydroxytoluene, nordihydroguaiaretic acidand propyl gallate.

[0040] Examples of chelating agents include sodium edetate and sodiumcitrate.

[0041] Examples of antibiotics include penicillin, neomycin,cephalothin, potassium permanganate, selenium sulfide, erythromycin,bacitracin, tethacyclin, chloramphenicol, vancomycin, nitrofurantoin,acrisorcin, chlorodontoin, and flucytosine.

[0042] Some of these additives function to enhance the efficacy of thecomposition by increasing the stability or percutaneous absorbability ofthe essential components of the present invention.

[0043] Also, any dosage form is acceptable, whether in solution,emulsion, powder dispersion, or others. Applicability is wide, includingfundamental dosage forms such as lotions, emulsions, creams and gels.

[0044] The composition of the present invention is preferably formulatedaccording to Formula 1:

[0045] Phenoxyethanol 1.0% (w/w)

[0046] Carbomer 0.5% (w/w)

[0047] Ovalbumin 20.0% (w/w)

[0048] Triethanolamine 0.3% (w/w)

[0049] Suitable vehicle 79.2% (w/w)

[0050] The method for processing and pasteurizing hen egg whites is wellknown in the art and generally involves reverse osmosis, heating, anddrying steps resulting in solid egg whites ready for compounding.

[0051] In addition to those stated above, suitable vehicles, carriersand adjuvants include water, Vaseline, petrolatum, mineral oil,vegetable oil, animal oil, organic and inorganic waxes, polymers such asxanthanes, gelatin, cellulose, collagen, starch, kaolin, carregeenan,gum arabic, synthetic polymers, alcohols, polyols, and the like. Thecarrier can also include sustained release carrier such as lypizomes,microsponges, microspheres, or microcapsules, aqueous base ointments,water in oil or oil in water emulsions, gels or the like.

[0052] The dose administered to an animal, particularly a human, in thecontext of the present invention should be sufficient to effect atherapeutic response over a reasonable time frame. The dose will bedetermined by the strength of the particular compositions employed andthe condition of the person. The size of the dose and the frequency ofapplication also will be determined by the existence, nature, and extentof any adverse side effects that may accompany the administration of aparticular composition.

[0053] The composition of the present invention may be employed to treatdiabetic ulcers, healing resistant wounds, bed sores, burns,osteomyelitis, trauma wounds, subcutaneous trauma and various forms ofdermatitis.

[0054] The following examples illustrate the inventive compositions andmethods, but should not be regarded as limiting the invention in anymanner. Additional studies are in progress as of the filing date of thisapplication.

EXAMPLE 1

[0055] In March 1998 a female patient who underwent laminectomy andfusion of T3 and T4 began applying the inventive composition to thesurgical wound. This patient experienced no scaring from the surgicalwounds and continued to be without scars 10 months post surgery. Thispatient had a previous history of radical surgery in May 1960 forremoval of a right breast tumor. The wound from that surgery healed inapproximately one month, but left a large scar. Eighteen months afterthe May 1960 surgery she had major abdominal surgery, and in 1985 hadreconstructive of the right and left breast, but with both surgeriesexperienced significant scaring. Thus, with the recent surgery, theinventive composition prevented scaring.

EXAMPLE 2

[0056] The wound healing characteristics of the inventive compositionwas demonstrated on Sprague-Dawley rats using a cream according toFormula 1 applied to a wound once daily for five days per week. Thepercentage of wound acceleration in days and in size was compared tocontrol. The percent of wound acceleration in days was determined to be5.19, and the percent of acceleration in size was determined to be22.07.

[0057] Note: Percentage wound acceleration in days=(1−TD/CD)×100%, whereTD and CD are the days required for 80% wound healing in the treatmentand control animals, respectively. Percent wound acceleration insize=(1−TA/CA)×100%, where TA and CA are the areas of the wounds in thetreatment in control animals, when 80% wound healing is reached on theformer.

EXAMPLE 3

[0058] A male patient routinely plagued by muscle and joint soreness andstiffness related to sports activities on the morning following suchactivities, applied the inventive composition to the affected areas andfound that he experienced no joint soreness the mornings following thesports activities.

EXAMPLE 4

[0059] Patient number 964885 has been applying the inventivecompositions to a diabetic ulcer on the left foot and has experienced ashrinkage of the wound.

EXAMPLE 5

[0060] A female applied the inventive composition in a cream form to athird degree burn on her left inner forearm. After daily application ofthe cream to the affected area, the blister reabsorbed readily, and thearea healed within seven days with no residual scar tissue.

EXAMPLE 6

[0061] An 88-year-old patient sustained a fall resulting in severalfacial abrasions and a 1.5 inch laceration over the right eyebrow. Withthe use of the inventive cream to the affected area, these lacerationshealed in about 72 hours leaving no scar tissue.

EXAMPLE 7

[0062] A male who has had diabetes for more than 30 years sustainedvarious skin tears and small ulcers on his upper and lower extremities.Daily application of the inventive composition in a cream form has keptthe affected areas clean and painfree and healing took place in arelatively short time frame.

EXAMPLE 8

[0063] A wound on a dog's hind leg was so extensive that the pad of thepaw was almost detached. The inventive composition cream was applied andthe wound bandaged. A few days later, the veterinarian noticedremarkable healing in that the wound was clean and dry with a goodgranulation bed, and there was the beginning of granulation across thewound. The extent of the healing was so great that the dog was able togo home with no bandage on its paw.

[0064] Similarly, a filly who had experienced a cut foreleg to thecannon bone was administered the inventive cream and the wound waswrapped. Two days later, upon removal of the bandage and checking of thewound, it had healed nicely across with a base of granulation over theperiosteum and showed a good, clean, dry wound bed with moist, suppleedges.

EXAMPLE 9

[0065] Several patients have used the inventive composition forarthritis, foot calluses, dry skin, back pain, bruises, allergies(including latex allergies), and inflammation. All such patientsreported symptomatic relief using the inventive composition.

[0066] Analysis of the Composition

[0067] The composition of the invention (formula 1) was tested usingantibodies directed against bovine and other mammalan derived growthfactors. In particular, the antibodies used were targeted toward PDGF,FGF a, FGF b, TGFβ, TGFα, and EGF. The following antibodies were usedfor detection of the growth factors. Unless noted otherwise theantibodies were purchased from Research Diagnostics Inc., Flanders, N.J.Antibodies Specificity Catalog Number Identification According to LabelRDI-PDGFABabg goat anti-PDGF Recognizes PDGF-AA, -AB, -BB chains ofhuman, primate, bovine and porcine. RDI-BFGFAabm mouse anti-FGFRecognizes bovine acidic Acidic FGF, human acidic FGF and human basicFGF. RDI-BRGFBabm2 mouse anti-bovine Recognizes bovine, rat, b. mouseand human FGF- FGF Basic RDI-TGFBabmx mouse anti-human Recognizes human,mouse TGF-B and bovine TGFB1 and B2, also Xenopus TGFB3. RDI-TGFAabmbmouse anti-human Recognizes mouse, rat and TGF-a human TCG-aRDI-MSEGFCabg goat anti mouse EGF Recognized mouse and C-term human EGFat carboxy terminus

[0068] Secondary Antibodies

[0069] From Jackson Laboratories:

[0070] Donkey anti-goat IgG (H&L) linked to biotin SP used for EGF andPDGF antibodies.

[0071] From Vectastain:

[0072] Horse anti-mouse (IgG (H&L) linked to biotin used for all otherantibodies.

[0073] Other Reagents

[0074] From Zymed:

[0075] Streptavidin linked to Horse Radish Peroxidise (HRP) (1:4000)

[0076] Protein Determination

[0077] Protein concentration was determined by the BCA assay (Pierce)according to manufacture's instructions.

[0078] Samples

[0079] Sample Provided by B&M Technologies: ppt MG54-2 — 21 mg/ml 1MG54-2 Jul. 24, 1997 115 mg/ml  2 MG54-3 Sep. 8, 1997 55 mg/ml

[0080] Crude Rat Brain Homogenate:

[0081] For control samples crude rat brain homogenates were generatedand run in adjacent lanes to samples of the inventive composition.

[0082] Western Blot

[0083] Separation of proteins was carried out in modified form from theprocedures of Laemmli (NATURE 227:680-685 (1970)). Samples and controlswere electrophoresed on a 15% SDS-PAGE gel. 40 μg of sample or-controlwas loaded onto each lane. Samples to test for PDGF and TGFB were rununder non-reducing conditions as specified by the supplier of theprimary antibody. All other samples and controls were run under reducingconditions. The samples were electrophoresed for 45 minutes at 200 V.

[0084] The gel was placed on a PVDF membrane and electrophorecticallytransferred for one hour at 25-30 amps. The membrane was blocked for onehour in PBS containing 5% nonfat dry milk. The primary antibodies werediluted in preparation for incubation with the membrane. Unlessotherwise noted the antibodies were diluted in PBS. The concentration ofthe antibodies used was as follows: Goat anti-Human PDGF-aB 10 ug/mlMouse anti-human TGFB  1 ug/ml in PBS with 1 mg/ml BSA Mouse anti-bovinebasic FGF1 ug/ml Mouse anti-bovine acidic FGF  1 ug/ml in PBS with 5mg/ml BSA Mouse anti-human TGF a 10 ug/ml Mouse anti-goat EGF 10 ug/ml

[0085] Lanes were cut from the membrane and incubated overnight at 4° C.with the primary antibody of interest. The membrane strips were washedfor one hour in PBS-tween (0.05% Tween 20) and incubated for two hourswith the secondary antibody of choice diluted in PBS. For landesincubated with goat derived primary antibodies, biotin linked donkeyanti-goat IgG at 1:10,000 dilution was used as secondary antibody. Forlanes incubated with mouse derived primary antibody, biotin linked horseanti-mouse IgG at 7.5 μg/ml diluted in PBS (1:200) with 1% normal horseserum was used as secondary antibody. The membrane strips were washedfor one our in PBS and incubated with either streptavidin-HRP for onehour (Zymed 1:4000, for donkey anti-goat IgG stained strips) orVectastain AB reagent (Avidin DH linked to Biotinylated peroxidase) for30 minutes (for horse anti-mouse IgG stained strips). The strips werewashed for 30 minutes in PBS and developed using diaminobenzadinetetrahydrochloride (DAB). The membrane strips were washed for 10 minutesin PBS and air dried.

[0086] Results

[0087] A preliminary Western run to test the response of the antibodieswas run without controls. Antibodies to PDGF and TGF β showed littleactivity against the inventive composition (lane 6 PDGF, 7 TGF β).Antibodies to EGF, TGF α, FGF α, FGF β, (lanes 1-4, respectively) hadidentical binding to the invention with bands at 70 kDa, 32-34 kDa and15-17 kDa.

[0088] While the specific invention has been described with an emphasisupon preferred embodiments, it will be obvious to those of ordinaryskill in the art that variations in the preferred methods of the presentinvention may be used and that it is intended that the invention may bepracticed otherwise than as specifically described herein. Accordingly,this invention includes all modifications encompassed within the spiritand scope of the invention as defined by the following claims.

I claim:
 1. A method for reducing or preventing the formation of scartissue in a subject comprising: applying a pharmacologically effectiveamount of a composition to the subject, the composition comprising about5.0% to 20% by weight crude ovalbumin containing a growth factorselected from the group consisting of epidermal growth factor,transforming growth factor alpha, fibroblast growth factor alpha, andfibroblast growth factor basic; about 1.0% by weight phenoxyethanol,about 0.5% by weight carbomer; and about 0.3% by weight triethanolamine;wherein the composition reduces or prevents the formation of scar tissuein the subject.
 2. The method of claim 1, wherein the growth factor isepidermal growth factor.
 3. The method of claim 1, wherein the growthfactor is transforming growth factor.
 4. The method of claim 1, whereinthe growth factor is fibroblast growth factor alpha.
 5. The method ofclaim 1, wherein the growth factor is fibroblast growth factor basic. 6.The method of claim 1, wherein the composition is in the form of atopical composition.
 7. The method of claim 1, wherein the compositionis in the form of a gel, cream, lotion, ointment or powder.
 8. A methodfor stimulating fibroblast production in a subject comprising: applyinga pharmacologically effective amount of a composition to the subject,the composition comprising about 5.0% to 20% by weight crude ovalbumincontaining a growth factor selected from the group consisting ofepidermal growth factor, transforming growth factor alpha, fibroblastgrowth factor alpha, and fibroblast growth factor basic; about 1.0% byweight phenoxyethanol, about 0.5% by weight carbomer; and about 0.3% byweight triethanolamine; wherein the composition stimulates fibroblastproduction in the subject.
 9. The method of claim 1, wherein the growthfactor is epidermal growth factor.
 10. The method of claim 1, whereinthe growth factor is transforming growth factor.
 11. The method of claimI, wherein the growth factor is fibroblast growth factor alpha.
 12. Themethod of claim 1, wherein the growth factor is fibroblast growth factorbasic.
 13. The method of claim 1, wherein the composition is in the formof a topical composition.
 14. The method of claim 1, wherein thecomposition is in the form of a gel, cream, lotion, ointment or powder.